This grant is requested to determine the molecular mechanisms of regulation of PLC delta1. The ability of proteins (such as G proteins), kinases, calcium fluxes, and lipids to regulate PLC delta1 activity will be assessed biochemically in vitro with purified components and biologically in vivo using transfeCted mammalian cells. Our work thus far has identified a regulatory domain in the N-terminal portion of a human PLC delta1 molecule. This region contains a recently identified 100 amino acid domain termed the pleckstrin- homology (PH) domain, which is present in many molecules involved in signal transduction. While several investigators have proposed that the PH domain is involved in mediating protein-protein interactions, our data instead suggest that this domain mediates lipid-protein interactions. We have demonstrated binding of several proteins containing PH domains (PLC delta1 and beta-adrenergic receptor kinase (beta-ARK)) to phospholipids with specificity and high affinity (mu M). Initial mutagenesis studies on PLC delta 1 have localized the phospholipid binding site to sub-domains 1 and 2 of the PH domain. We will test the hypothesis that PLC delta1 is primarily regulated by the translocation of the effector enzyme to its substrate phospholipid through the PH domain.